The modulation of neuronal function by various kinds of endogenous opioid peptides is one of the most important cellular events of brain function. However, not much is known about the cellular and membranous mechanisms of opioid actions. The objective of this grant proposal is to elucidate the ionic channel and second messenger mechanisms of two kinds of endogenous opioid peptides, beta-endorphin and dynorphin. Primary cultures from two brain regions of postnatal animals, the locus coeruleus and the substantia nigra will be used. Electrophysiological techniques including the whole cell version of patch clamp and intracellular microelectrode methods will be used. The proposal is divided into two projects. The first project deals with the effects of beta-endorphin on dissociated cultured noradrenergic neurons from the locus coeruleus. A preliminary study in the laboratory shows that beta- endorphin produces a slow hyperpolarization at the resting stare, resulting in an inhibition of neuronal activity. The preliminary study also shows that pertussis toxin inhibits this hyperpolarization, suggesting that inhibitory GTP-binding proteins may be involved as second messengers. To investigate further the precise ionic and second messenger mechanisms of this slow hyperpolarization is the aim of the first project. The second project consists of two parts. The first part is to make primary dissociated cell culture of the substantia nigra of postnatal rats and mice. Dopaminergic neurons will be identified by using immunohistochemical or cathecholamine histofluorescence methods. The second part is to investigate the effects of dynorphin on cultured neurons from the substantia nigra, which is known to be richly innervated with dynorphin-containing nerve fibers. The ionic and second messenger mechanisms of dynorphin effects will be investigated.